Recent unpublished observations have established the predicted capacity of frozen-thawed mouse spermatozoa to fertilize and to induce the development of normal progeny. However, the low number of surviving motile spermatozoa is insufficient to realize the potential benefits of cryobanking in mouse breeding, especially since they show reduced fertility and induce the production of smaller litters. In contrast, cryopreserved bovine semen with significant percentages of moving cells can be extended significantly for successful mission-directed use in multiple inseminations, yielding many progeny per ejaculate and undiminished rates of fertility. The potential for such practical cryobanking has been demonstrated recently with rabbit spermatozoa. The objective of this project is to devise a method of cyropreservation for mouse spermatozoa which will increase the number of motile and reproductively functional cells to a level commensurate with practical cryobanking as required in controlled genetic breeding. Factors of cryosurvival to be evaluated in the formulation of such a method will include: extension of spermatozoa with various media; pretreatment with cryoprotective agents as to amount, cellular location and toxicity; sensitivity to and protection from temperature shock; rates of cooling and rewarming during intervals on a temperature change curve from above freezing to -19 degress C and back; frozen storage containers and conditions; and latent cryoinjury. Cryosurvival will be assessed in terms of percent and type of motility, membrane integrity, in vitro fertilization and in vivo insemination with progeny production.